This invention relates to DNA and peptide sequences encoding a novel mammalian secreted group IIF phospholipase A2 and more particularly, a novel human group IIF phospholipase A2. The invention also relates to the use of this enzyme in methods for screening various chemical compounds.
Phospholipases A2 (PLA2, EC 3.1.1.4.) form a superfamily of enzymes that catalyze the hydrolysis of glycerophospholipids at the sn-2 position, producing free fatty acids and lysophospholipids [1-4]. Many intracellular and secreted phospholipases A2 (sPLA2s) have been cloned in recent years [2,5], and several of them are involved in a variety of physiological and pathological functions including lipid digestion, cell proliferation, production of lipid mediators of inflammation, antibacterial defense, and cancer [4,6].
Within the phospholipase A2 superfamily, sPLA2s form a relatively homogenous family of enzymes. They are characterized by the presence of several disulfides, an overall conserved three-dimensional structure and a common Ca2+-dependent catalytic mechanism. Five novel mouse sPLA2s have been cloned during the last three years [7,8], and the mouse sPLA2 family now comprises 8 distinct 14-16 kDa sPLA2s called group IB, IIA, IIC, IID, IIE, IIF, V and X, as well as otoconin-95, a sPLA2-like protein with peculiar structural properties [9,10]. Interestingly, genes for group IIA, IIC, IID, IIE, IIF, and V sPLA2s all map to mouse chromosome 4, suggesting the existence of a sPLA2 gene cluster on this chromosome [8]. Group IB, IIA, IID, IIE, V and X sPLA2s, but not group IIF have been cloned from humans [11-13]. Conversely, group IIC sPLA2 appears as a pseudogene in humans [14].
In addition, a novel human sPLA2 with a predicted molecular mass of 55 kDA and a central domain similar to insect group III sPLA2s has recently been cloned [15], but it remains to be determined if this sPLA2 is functional in the mouse species. This novel human sPLA2 is also disclosed in the French International Patent Application No01/59129. All mouse and human sPLA2s have distinct tissue distributions, suggesting that each of them exert non redundant functions that could be related to their different enzymatic properties [6,16,17], and/or their binding properties to specific receptors [17-19].
A comprehensive abbreviation system for the various sPLA2s is used thereafter: each sPLA2 is abbreviated with a lowercase letter indicating the sPLA2 species (m, h, for mouse and human, respectively) followed by capital characters identifying the sPLA2 group (GI, GII, GIII, GV, and GX) and subgroup (A, B, C, D, E, F).
This invention relates to a mammalian secreted group IIF sPLA2 which is Ca2+-dependent, maximally active at pH of about 7-8, and hydrolyzes phosphatidylglycerol versus phosphatidylcholine with about a 15-fold preference.